How is it really working - The smallest locial unit - establishing an equipollent to an electronic

transistor unit

THe main goal is to provide universality, so input and output on same basis, RNA, transcribiton, sin

THe main goal is to provide

switch

• switching element
  

The switching unit is the functional core element of our switches, allowing a shift between an "on" and "off" state. Since we work on the level of transcription, a "switchable" transcriptional terminator is suitalbe for this purpose.

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The special thing about it is all of our switches consist of THE SAME switching unit, so having found one functional "switchable" terminator will allow almost unlimited upscaling. This is the main difference to previous works on this field, which always required to develop a new shifting principle for each switch. Beside the extendablity furhter advantages are we can provide a comparable on/off shifting rate. This makes our switches more similar to transistors than ...

Practically, the shift between the two states is induced by complementrary RNA-sequences, influencing the terminators secoundary structure. To seperate

this switching element from the specific accessiblity, we designed a synthetic switching unit relying on NUPACK simulations, which is not able to change the terminators state on its own, but only in combination with a specific recognition site:

• recognition site
  

It defines the specific access of one of our switches by an input molecule. Therefore, a unique

recognition element is assigned to each switch. This allows to to arrange and interconnect numerous

of these switches in a specif logical order, comparable to wires connecting different transistors.

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signals, the inputs and outputs of the bioLOGICS switch

Signals present the "trigger" to shift switches between on and off. It requires the ability to change the terminators secondary structure BUT, only if a special recognition site is detected.

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The bioLOGIC switch with regard of logical operations

As described, each switch can be accessed by a specific RNA-signal molecule, illustrating the input. In turn, another RNA-signal molecule will be produced if the switch shifts its state, now beeing the output of one switch and at the same time, a possible input for the following switch or several ones. This easily allows arranging several switches in specific sequences and faulty wiring - the corner stone of a logical network.

To easen the building of logical networks, applying mathematical logcis, e.g. Boolean logics like in computaltion science would be worthwile. It is possible to establish general Boolean operators with our switches and thus build "logical modules".

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Evaluation

To evaluate functionality of our molecular switches, we developed several in vivo and in vitro assays and relied on existing assays. Thus we don't want to develop additional biobricks, we rather want to establish a platform to realize the full opportunity of the biobrick system. This would enable people easily setting up sensor - reporter circuits AND interconnect them to complete biological chips... the way to real artificial cells and synthetic biology. (zu krass?)

Network construction

Designing complex biological networks based on either traditional protein engineering or our new bioLOGICS is still a complex task. We developed a software which allows the fast construction of a bioLOGICS based networks.
To read more about this, look at our Software page

Our switches are based on the principle of transcriptional antitermination. Transcription can be cancelled by the formation of a RNA stem loop in the nascent RNA-chain, which then causes the RNA polymerase to stop transcription and fall off the DNA. We use sequences as transcriptional switches, that are calculated to be capable of stem loop formation.

Novel switches based on RNA-RNA interaction

We plan to control the termination at these switches with a small RNA molecule (our RNA "signal") that is complementary to a part of the stem loop forming sequence. This small functional RNA inhibits the stem loop formation by complementary base-pairing and hence avoids transcription termination. The signal is composed of two parts: While the first part provides specifity (recognition site), the second part causing stem loop disintegration (functional core) can in principle be the same for all bioLOGICS. Therefore variation of the first part allows the construction of an endless number of switches. The functional core causing stem loop disintegration is based on a working system (see attenuation) established by nature. Different stem loops were tested in this effort: Regulatory parts from the E. coli trp-operon, his-operon and one based on previous iGEM-work.
The initial signal can be provided by various metabolic compounds and stimuli and the output signal can be anything DNA-coded, too. In the last years, many working sensory systems were submitted to the Partsregistry. Those parts can now be utilized as inputs for our network. BioLOGICS provides a new way to use the whole potential of iGEM distributions connecting different parts for totally new applications.
The major advantage over conventional protein based devices and even riboswitches is the small size of our basic units, its easy construction, huge variability and easy upscaling to complex networks in cells. Furthermore introduction of an RNA network into E. coli is especially easy - all informations can be coded on one plasmid. In comparison to protein networks, the elegance of RNA based networks lies in its functionality, simplicity and predictability. Since RNA-RNA interactions are highly predictable and since our system is based on the variation of one principle in many switches, simulations of our networks are much more accurate than those of comparable systems currently available. Incorporation of established Biobricks allows a high diversity of input and output signals providing a whole new level of cell control.